FIELD OF THE INVENTION
The initial stages of the biological revolution demonstrated the feasibility of obtaining expression of mammalian genes in lower organisms. Because of the much greater amount of knowledge associated with the regulatory sequences of bacteria, bacteria were chosen as the initial host for producing heterologous proteins. However, bacteria have many shortcomings. Not least of these shortcomings is the fact that they produce an enterotoxin which must be completely removed, if the product is to be administered to a mammalian recipient, e.g. as a pharmaceutical agent.
Furthermore, the codons of the heterologous genes will be expressed with relatively low efficiency, since the preferred codons of the source of the heterologous protein and the host will be substantially different. In addition, where the product of interest needs to be processed, such as glycosylated, matured by removal of polypeptide sequences, or assembled, bacteria frequently prove to be incapable or inefficient at these processes. Moreover, for commerical application of genetic engineering technology it would be desirable for ease of subsequent purification if synthesized products were secreted into the growth medium, a process in bacteria of only limited, laboratory scale use. It is therefore desirable to find alternative hosts.
Yeast as a host has many advantages which recommend its use. The commercial fermentation of yeast is well established. Yeast is a eukaryote unlike bacteria, so that it shares greater similarities with mammalian organisms. Yeast are thus capable of many of the processing steps observed in higher organisms and secretion of several natural polypeptides and proteins is known. Furthermore, yeasts do not produce enterotoxins.
It is therefore desirable to provide yeast regulatory signals which may be employed for the efficient production of heterologous proteins in yeast. While the existence of the regulatory signals may be predicted, their isolation, manipulation, and ultimately establishing that the regulatory signals can operate with alien flanking regions in conjunction with a foreign DNA sequence is long and arduous work, requiring well thought out experimental design, careful manipulation, and rigorous proofs of having achieved the intended result at each of the many stages involved.